1. Field of the Invention
The present invention relates generally to the fields of molecular biology, cardiovascular medicine and transgenic animals. More specifically, the present invention relates to a novel transgenic mouse model of atherosclerosis.
2. Description of the Related Art
Although atherogenesis involves many different events, abnormal accumulation of cholesterol in the artery wall has long been considered an essential element of the atherogenic process. Cholesterol deposits in the atherosclerosclerotic plaque represent both intracellular lipid deposits (foam cells) and extracellular lipid particles. The foam cells are caused by the abnormal uptake of certain lipoprotein species in circulating blood by macrophages. The origins of extracellular lipid particles are either infiltered plasma lipoproteins or cellular necrotic products.
Several reports indicate that human atherosclerosis may be a reversible process (1). Regression of experimentally induced atherosclerosis in experimental animals has been reported following dietary manipulation (2) or by the administration of hypolipedemic agents (2). Prospective epidemiological studies indicate that a strong positive correlation exists between plasma levels of LDL and VLDL. VLDL and LDL components have also been identified in human atherosclerotic plaques. However, normalipedemic patients also develop premature coronary artery disease. It has been suggested that elevated levels of HDL inhibits the development of atherosclerosis by acting as a sink for cholesterol from foam cell membranes.
It has been postulated that cholesterol lowering stabilizes atherosclerotic plaques, particularly those with eccentric, lipid-rich cores. Lesions with lipid-rich cores are particularly susceptible to intravascular hemorrhage, and are usually associated with clinical events. Foam cells are often found at the sites of plaque rupture, suggesting that plaque erosion by foam cells may be a major mechanism in predisposing atherosclerosis (12). A reduction in lipid content may alter the structure of these plaques in such a way that they are less vulnerable to rupture and other complications. This theory is in agreement with the concept of reverse cholesterol transport. In this theory, high levels of HDL are thought to promote cholesterol efflux from cells. It is therefore possible that, while reduced levels of LDL inhibit further lipid accumulation in lesions, increasing levels of HDL by 1% decreases coronary artery disease mortality by three percent.
Lipoproteins are macromolecular assemblies of specific apolipoproteins and lipids. The lipids are associated with protein components by noncovalent forces. All of the lipoproteins have cores of triglycerides and cholesteryl esters with the surface stabilized by phospholipids, cholesterol and apolipoproteins. There are some HDL particles that are devoid of core. These are discoidal and nascent particles and the newly classified pre.beta.HDL particles. During the hydrolysis of chylomicrons and VLDL some surface remnants are formed which are vesicular and these vesicular substances are associated with exchangeable apolipoproteins.
High Density Lipoproteins
Recent studies provide a strong support for the role of plasma lipoproteins in atherosclerotic plaque formation. Plaque stabilization due to rapid removal of cholesterol from macrophage-derived foam cells has been suggested to explain the rapid cessation of clinical coronary artery disease events. Epidemiological studies show an inverse correlation of HDL and apo A-I to atherosclerosis.
The inverse relationship between atherosclerosis and HDL has been well established. One prominent role of HDL appears to be that HDL is involved in the reverse-cholesterol transport, i.e., the transportation of cholesterol from peripheral tissue to the liver (3). HDL may have other beneficial roles. HDL, by regulating the inflammatory response, may also regulate the tissue injury seen in inflammatory disorders. HDL decreases antibody production and inhibits lymphocyte cellular cytotoxicity. There is a pool of apoA-I free from HDL which has been shown to effectively efflux cellular cholesterol.
Recently, it has been shown that HDL delivers cholesterol to the cells through the class B scavenger receptor SRBI. This receptor binds HDL with high affinity, and is primarily expressed in the liver and nonplacental steroidogenic tissues. Thus, HDL metabolism is distinct from LDL metabolism in which cholesterol alone is taken up by the cells and not apoA-I.
Low Density Lipoproteins
High levels of LDL are the main cause of hypercholesterolemia. LDL are enriched with free cholesterol. Due to its size (21 to 25 nm), LDL can be filtered into the arterial wall where they can initiate the atherogenic process. LDL is derived from the catabolism of VLDL. This indicates that any alteration of VLDL catabolism should alter the production of LDL. The precise mechanism whereby VLDL remnants are converted to LDL is not known.
LDL is cleared via the LDL receptor. During this process, the entire LDL particle is endocytosed, the protein apoB is proteolyzed and the cholesterol is taken up by the cells. There are other nonreceptor-dependent pathways whereby LDL is removed. Accumulating evidence indicates that oxidized LDL is atherogenic. In vitro evidence suggests that HDL is an antioxidant and prevents self-aggregation. These latter effects are thought to be directly related to the apoA-I component present in HDL.
Very Low Density Lipoproteins
Very Low Density lipoproteins are also called the Triglycerides (TG)-rich lipoproteins. Triglycerides in VLDL are hydrolyzed by the action of lipoprotein lipase (LPL). ApoC-II, an activator of LPL is present on the surface of VLDL. This hydrolysis produces surface-remnants enriched with phospholipids and core remnants. The smaller core remnants are ultimately converted to LDL. The exchangeable apolipoproteins present on the surface of VLDL are responsible for the function of VLDL. In LDL-receptor deficient mice fed a high fat diet, overexpression of LPL inhibited diet-induced atherosclerosclerosis. Thus, increased catabolism of VLDL can inhibit atherosclerosis despite the fact the cholesterol is not metabolized via the VLDL.
Exchangeable Apolipoproteins
One of the main functions of exchangeable apolipoproteins appears to be to solubilize otherwise insoluble lipids in circulating plasma. The mechanism by which the exchangeable apolipoproteins associate with phospholipids was first proposed by Segrest et al. This group of proteins interacts with lipids through specialized helical domains (the "amphipathic helical domain". There is experimental evidence that the amphipathic helical domains are also important for functional properties of lipoproteins. ApoA-I is the major activator of the plasma enzyme LCAT that converts free cholesterol into cholesteryl ester. ApoA-I is also capable of effluxing cholesterol from cholesterol loaded cells. This may have a direct implication on the ability of HDL and apoA-I to inhibit atherosclerosis. In addition, ApoE, the protein component of VLDL, is involved in the receptor-mediated removal of VLDL.
Animal Models for Atherosclerosis
The genetically inbred mouse has been useful in deciphering the impact of a specific gene products on the lipoprotein metabolism. Currently, murine models of atherosclerosis have been used entensively for the study of atherogenic lipoproteins. However, there exists a large variation in the susceptibility of diet-induced atherosclerosis across the strains. C57BL/6 mice, the most susceptible strain, shows the largest reduction of HDL cholesterol with high fat diet administration. This implies that HDL is the major determinant of atherogenicity.
Mice overexpressing human apoA-I are resistant to diet-induced atherosclerosis. Mice overexpressing apoA-II or CETP developed severe diet-administered atherosclerosis. Moreover, mice lacking apoE develop atherosclerosis spontaneously, even without high fat diet administration. Similarly, LDL receptor gene knockout mice also develop atherosclerosis and are considered to be models of familial hypercholesterolemia. These produce severe atherosclerosis when fed with a high fat diet.
Other animal models, such as rabbits, can be used for studying the role of apolipoproteins in atherosclerosis. However, it is difficult to genetically breed to produce transgenes in other animals. Another drawback in the transgenic mouse models of the prior art is due to the size of the animals, it is difficult to obtain several samples of plasma during a study to measure changes in lipoprotein profiles that occur during an experiment.
The prior art is deficient in the lack of a novel transgenic mouse model of atherosclerosis. The present invention fulfills this longstanding need and desire in the art.